Identification of RNA partners of viral proteins in infected cells

被引:13
|
作者
Komarova, Anastassia V. [1 ]
Combredet, Chantal [1 ]
Sismeiro, Odile [2 ]
Dillies, Marie-Agnes [2 ]
Jagla, Bernd [2 ]
David, Raul Yusef Sanchez [1 ]
Vabret, Nicolas [1 ]
Coppee, Jean-Yves [2 ]
Vidalain, Pierre-Olivier [1 ]
Tangy, Frederic [1 ]
机构
[1] Inst Pasteur, CNRS, URA 3015, Unite Genom Virale & Vaccinat, Paris, France
[2] Inst Pasteur, Plateforme Transcriptome & Epigenome Genopole, Paris, France
关键词
RNA-protein interactions; next-generation sequencing; affinity purification; tagged proteins; measles virus; nucleocapsids; DEFECTIVE INTERFERING PARTICLES; DIFFERENTIAL EXPRESSION ANALYSIS; MEASLES-VIRUS; MESSENGER-RNA; VACCINE STRAINS; RIG-I; GENERATION; PURIFICATION; REPLICATION; GENOME;
D O I
10.4161/rna.24453
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
RNA viruses exhibit small-sized genomes encoding few proteins, but still establish complex networks of protein-protein and RNA-protein interactions within a cell to achieve efficient replication and spreading. Deciphering these interactions is essential to reach a comprehensive understanding of the viral infection process. To study RNA-protein complexes directly in infected cells, we developed a new approach based on recombinant viruses expressing tagged viral proteins that were purified together with their specific RNA partners. High-throughput sequencing was then used to identify these RNA molecules. As a proof of principle, this method was applied to measles virus nucleoprotein (MV-N). It revealed that in addition to full-length genomes, MV-N specifically interacted with a unique population of 5 copy-back defective interfering RNA genomes that we characterized. Such RNA molecules were able to induce strong activation of interferon-stimulated response element promoter preferentially via the cytoplasmic pattern recognition receptor RIG-I protein, demonstrating their biological functionality. Thus, this method provides a new platform to explore biologically active RNA-protein networks that viruses establish within infected cells.
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页码:944 / 957
页数:14
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